On 25 August 2007, Puerta collapsed and lost consciousness in the penalty area due to a cardiac arrest during Sevilla's first La Liga match of the 2007–08 campaign at homeground Sánchez Pizjuán against Getafe CF.[7] He was seen crouching and then subsequently collapsing upon moving back to his team's goal after only 35 minutes of the game had passed,[3] as teammates Ivica Dragutinović and Andrés Palop immediately ran to his side as he lost consciousness; moments later, club medical staff and other players followed suit.

After recovering and being substituted, Puerta was able to walk to the dressing room, where he collapsed once again. He was resuscitated by the doctors and taken, by ambulance, to the intensive care unit of Virgen del Rocío hospital, where he received cardiopulmonary resuscitation.

As a mark of respect, players from both Sevilla and city rivals Real Betis attended his funeral days after his death. Following Puerta's death, FIFA ordered the installation of resuscitation rooms in every stadium that hosted the World Cup qualifiers.

Additionally, Sevilla's UEFA Super Cupgame with A.C. Milan on 31 August went ahead, as a tribute to the player, with all 22 participants having the name 'PUERTA' printed on the back of their jersey. The players and officials on both sides also wore black armbands.[12]

Sevilla subsequently retired Puerta's number 16 shirt, with the provision that should his son, Aitor Antonio (born 22 October 2007),[13] one day play for the club, he will have the option to bring the number out of retirement.[14] However, Spanish football teams were not allowed to do so, as the Royal Spanish Football Federation stated that clubs should use number 1 to 25 for their regular squad, with no additional room for manoeuvre. As a result, David Prieto wore the shirt in 2007–08 in honour of his friend[15] but it was later decided that only youth products could wear that jersey.[16] Controversially, the number was given to the Argentine Federico Fazio in 2016.[17]

Ramos wore t-shirts in memory of Puerta after Spain won UEFA Euro 2008 and the 2010 FIFA World Cup, with Navas doing the same on the latter occasion.[18] Earlier in 2010, a statue of him was built in the club's José Ramón Cisneros Palacios sporting facilities.[19][20]